ABSTRACT Diseases of the nodal tissue of the heart can be life threatening, particularly in the young. Nodal tissue spontaneously depolarizes serving as a pacemaker for cardiac contraction, yet despite this central role critical for survival, the cause of nodal dysfunction is poorly understood. This lack of mechanistic understanding has impaired development of efficacious and selective pharmacotherapies, and as a correlate, drugs levied against nodal disease carry significant toxicity for the patient and can still be entirely ineffective. While the nodal automaticity was traditionally thought to be controlled by ion channels on the plasma membrane, there is a growing body of evidence that calcium-signaling within the cell may regulate spontaneous depolarization ? its automaticity. Previous investigation has shown that calcium leak from the internal calcium release channel, RyR2, may be associated with increased nodal firing, thus understanding this so-called ?calcium clock? can provide additional molecular targets for nodal-specific novel therapeutics. I have created a mouse model of nodal-specific expression silencing of a protein called junctophilin-2 (JPH2), which I have previously shown to be critical to effective calcium- handling in the contractile myocyte, as a tool for studying a dysfunctional calcium clock. I have found that this mouse has an elevated heart rate at rest and a rapidly firing atrioventricular node which causes an arrhythmia known as accelerated junctional rhythm (AJR). I propose 3 specific aims to test our central hypothesis that reduced JPH2 expression results in CaMKII-mediated increase in RyR2 gating which causes increased store calcium leak and drives increased nodal automaticity and AJR. My aims are to 1) utilize confocal-based calcium imaging of isolated nodal cells from HCN4:shJPH2 mice, coupled with RyR2 single channel recordings to determine whether reduced JPH2 expression causes increased calcium leak with higher RyR2 channel opening probability; 2) apply known chemical inhibitors of RyR2 to isolated single cells and HCN4:shJPh2 mice to assess whether calcium leak can be normalized and AJR effectively treated; and 3) conduct biochemistry from isolated nodal tissue to determine the role of CaMKII signaling, including its downstream phosphorylation targets, in regulation of nodal firing. I expect that completion of these aims will yield clinically translatable mechanistic insight into the ?calcium clock? of the node. Through exploration of the first murine model of isolated cardiac nodal disease in the literature, these aims will provide a substrate from which to test novel therapeutic agents specifically targeted at perturbed calcium-signaling in nodal tissue. Completion of this 5-year training grant will allow me to combine my clinical training in pediatric electrophysiology with exploration of the molecular mechanisms of nodal disease and become an independently funded physician-scientist committed to helping children with arrhythmias.